Protein degradation systems have revealed that catabolism is energy dependent. Inasmuch as proteolysis itself is not energetically unfavorable the immediate goal of the proposed research is to study this interesting paradox. A role for ATP in protein degradation has been implicated by others and lysosomes do have a ATPase. Since ATPase offers a direct link to cellular energy it will be given first priority. Specific antisera to rat-liver lysosomal ATPase will be used in suitable protein degradation systems to delineate the function of the ATPase in catabolism. The long range goals of the proposed research are (1) to determine the enzymes which facilitate membrane fusion and proteolysis, (2) to define the dependency of these reactions on cellular energy, and (3) to elucidate at the molecular level the regulatory mechanism by which individual proteins are degraded at different rates. Preexisting systems will be modified and, if necessary, new ones developed to study the energy-dependency of membrane fusion and proteolysis. An in vitro system involving the incubation of liposomes containing albumin with purifid lysosomes may encompass the advantages of previous systems and offer the new possibility of studying membrane fusion directly. After purification and characterization of the lysosomal ATPase, antisera will be prepared by immunization of rabbits and will be used as specific probes in test systems to investigate the energy requiring steps in the degradation process. The role of phospholipase and lysolecithin acylase activities in membrane fusion may also be examined in test systems using antisera as specific probes. In addition, antisera to the lysosomal ATPase will be used to measure the turnover rate of lysosomes by selective immunoprecipitation of the ATPase at various times after administration of 3-H-leucine.